Inhibition Effects of Essential Oil of Cymbocarpum

MAKÜ FEBED
ISSN Online: 1309-2243
http://febed.mehmetakif.edu.tr
Mehmet Akif Ersoy Üniversitesi Fen Bilimleri Enstitüsü Dergisi 3 (1): 32-36 (2012)
Research Paper / Araştırma Makalesi
Inhibition Effects of Essential Oil of Cymbocarpum erythraeum (Dc.)
Boiss., on Percentage of Survival from Larvae to Adult in Drosophila
melanogaster and its Chemical Composition
1
1
2
3
4
Özkan Aksakal , Handan Uysal , Deniz Altun Çolak , Ebru Mete , Yusuf Kaya
1
Department of Biology, Science Faculty, Atatürk University, 25240 Erzurum, Turkey
Department of Biology, Arts and Science Faculty, Erzincan University, 24100 Erzincan, Turkey
3
Department of Chemistry, Science Faculty, Atatürk University, 25240 Erzurum, Turkey
4
Department of Biology, Science Faculty, Atatürk University, 25240 Erzurum, Turkey
2
Received (Geliş Tarihi): 04.01.2012, Accepted (Kabul Tarihi): 29.03.2012
Corresponding author (Yazışmalardan Sorumlu Yazar): [email protected] (H. Uysal)
0 442 231 43 36
0 442 236 09 48
ABSTRACT
The essential oil of aerial parts of Cymbocarpum erythraeum (Apiaceae), collected in Palandöken Mountain in
Erzurum, Turkey, were obtained by hydro-distillation and analyzed by gas chromatography-mass spectrometry (GCMS). Out of 25 peaks 22 components, which constitute 96.36% were identified in the oil. The main constituents were
(E)-2-decenal, (2E)-dodecenal, 8S,14-cedranediol, n-tetradecanol and (E)-4-decenal. In this study, six different
concentrations of essential oil (15, 30, 60, 90, 120 and 150µL) were investigated for larvicidal effects on Drosophila
melanogaster. The essential oil of the aerial parts exhibited potent activity against third instar larvae of Drosophila
melanogaster with 15 to 150 µL. These properties suggest that essential oil of Cymbocarpum eryhhraeum is a
potential source of valuable larvicidal compounds for direct use as natural insecticides.
Key Words: Cymbocarpum erythraeum, Drosophila melanogaster, Essential oil, Larvicidal effect
Drosophila melanogaster’de Larvadan Ergine Hayatta Kalış Oranı Üzerine Cymbocarpum
erythraeum (Dc.) Boiss., Bitkisine Ait Esansiyal Yağın İnhibisyon Etkisi ve Kimyasal
Kompozisyonu
ÖZET
Palandöken Dağı’ndan (Erzurum/Türkiye) toplanan Cymbocarpum erythraeum’un (Apiaceae) toprak üstü kısımlarına
ait esansiyal yağ, su distilasyon yöntemiyle elde edildi ve kütle gaz kromatografi (GC-MS) yöntemiyle içeriği belirlendi.
C. erythraeum’a ait esansiyal yağı oluşturan toplam 25 bileşenin 22’si (%96,36) tanımlandı. Ana bileşenlerin (E)-2decenal, (2E)-dodecenal, 8S,14-cedranediol, n-tetradecanol ve (E)-4-decenal olduğu tespit edildi. Bu çalışmada
esansiyal yağın farklı konsantrasyonlarının (15, 30, 60, 90, 120 ve 150 µL) Drosophila melanogaster üzerindeki
larvisidal etkisi araştırıldı. Esansiyal yağın 15-150 µL arasındaki tüm konsantrasyonlarda güçlü larvisidal etki
gösterdiği belirlendi. Bu sonuçlara göre, C. eryhhraeum bitkisine ait esansiyal yağın doğal insektisit olarak direct
kullanımda değerli bir kaynak olduğu söylenebilir.
Anahtar Kelimeler: Cymbocarpum erythraeum, Drosophila melanogaster, Uçucu yağ, Larvasidal etki
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Ö. Aksakal, H. Uysal, D.A. Çolak, E. Mete, Y. Kaya MAKÜ FEBED 3(1): 32-36 (2012)
1. INTRODUCTION
Fly cultures and crosses were grown on standard fly
medium. The standard medium was a dried yeast-agarsugar medium (SDM). All experiments were carried out
at 25±1°C, under constant dark condition and 40-60%
relatively humidity. Prior to the experiments the stocks
had been maintained for many years in the laboratory of
the Department of Biology of the Atatürk University in
Erzurum was, therefore, highly inbred with little genetic
variation. The females used in this experiment were
virgins.
Essential oils are extracted from various aromatic plants
generally localized in temperate to warm countries like
Mediterranean and tropical countries where they
represent an important part of the traditional
pharmacopoeia (Bakkali, 2008). They are naturally
occurring substances which are often responsible for a
plant’s distinctive scent or taste. They can be
synthesized by all plant organs, i.e. buds, flowers,
leaves, stems, twigs, seeds, fruits, roots, wood or bark,
and are stored in secretory cells, cavities, canals,
epidermic cells or glandular trichomes (Bakkali, 2008).
Eggs from the crosses between virgin females and
males were collected during 4 h periods in culture
bottles containing SDM. After 72±4 hours larvae were
washed and selected for the treatment. For the chronic
feeding study, small plastic vials were prepared with 1.5
g dry Drosophila Instant Medium (Carolina Biological
Supply Company Burlington, NC, USA) and 5 mL of the
respective essential oil of Cymbocarpum erythraeum
(CEEO). SDM was prepared with distilled water for the
controls. 100 larvae were embedded into this medium.
The larvae were fed with different concentrations of the
extract of Cymbocarpum erythraeum. Feeding ended
with pupation of the surviving larvae. The experiments
were repeated three times. Concentrations of the
essential oil were 15 µL to 150 µL. After developmental
time, all surviving flies were scored respective of sex
and recorded.
During the last years, plant essential oils, plant extracts
and their constituents have received more and more
attention as potentially useful bioactive compounds with
particular emphasis on their insecticidal, larvicidal and
adulticidal activities (Miyazawa et al., 2004; Tsukamoto
et al., 2005).
Essential oils composed by isoprenoid compounds,
mainly mono- and sequiterpenes are the carriers of the
smell found in aromatic plants. The essential oils are
playing an important role in the interactions between
plants and insects. In particular, monoterpenoids,
components of essential oils in many plants, are very
important to insects because they can attract beneficial
insects, which can aid in pollination, and they can help
plants defend against harmful insects because of its
high influence with volatility (Grodnitzky & Coats, 2002;
Tsukamoto et al., 2005). Since these compounds often
biodegrade to nontoxic products, they could be much
safer insect control agents and more suitable for use in
integrated pest management (Dev & Koul, 1997;
Grodnitzky & Coats, 2002).
Hydro distillation was used for 4 hour to extract the
essential oils using a Clevenger-type apparatus. The
obtained essential oil was dried over anhydrous sodium
sulfate and, after filtration, stored at +4ºC until tested
and analyzed. The analysis of the essential oil was
performed using a Thermofinnigan Trace GC/Trace
DSQ /A1300, (E.I Quadra pole) equipped with a SGEBPX5 MS capillary column (30 m × 0.25 mm i.d., 0.25
µm). For GC-MS detection an electron ionization system
with ionization energy of 70 eV was used. Helium was
the carrier gas, at a flow rate of 1mL/min. Injector and
MS transfer line temperatures were set at 220°C and
290°C, respectively. The program used was 50–150°C
at a rate of 3°C/min, held isothermal for 10 minutes and
finally raised to 250°C at 10°C/min. Diluted samples
(1/100, v/v, in methylene chloride) of 1.0 mL were
injected manually and in the split less mode. The
components were identified based on the comparison of
their relative retention time and mass spectra with those
of standards, Wiley7N, TRLIB library data of the GC-MS
system and literature data (Adams, 2001). The
quantitative data were expressed as area %.
The characteristic features of Cymbocarpum erythraeum
are that low, glabrous annual or biennial; stems 10-30
cm, often branched from base. Basal leaves soon
withering; petiole long abruptly dilated into sheath,
lamina 10-15x10-15 mm, bipinnate, ultimate segments
3-8x0.5 mm, linear, acute. Cauline leaves similar but
smaller, subtended by expanded membranousmargined sheath. Umbel rays 9-11, unequal, glabrous.
Bracts and bracteoles 5 linear, acute, recurved, not
marginated. Flowers white (rarely pink); petals c. 1 mm.
Fruit 3-3.5x2.5 mm, ovoid, glabrous, ridge filiform,
slightly dilated at margins (Davis, 1988).
According to our best knowledge, there is no report on
the species of aromatic Cymbocarpum erythraeum.
Thus, the main aim of this study was to determine the
main constituents of the essential oil of Cymbocarpum
erythraeum aerial parts growing in Turkey and to
evaluate its larvicidal and adulticidal activity against
Drosophila melanogaster.
3. RESULTS and DISCUSSION
The essential oil of Cymbocarpum erythraeum obtained
on
hydro-distillation
was
analyzed
by
gas
chromatography-mass spectroscopy (GC-MS). Twenty
three components representing 96.36% of the total oil
were identified. The constituents identified by GC-MS
analysis, their retention indices and area percentages
(concentrations) are summarized in Table 1. As can be
seen from this Table, the major constituents in the
essential oil of Cymbocarpum erythraeum collected from
2. MATERIAL and METHODS
Cymbocarpum erythraeum aerial parts were collected
from Palandöken Mountain in Erzurum during flowering
time at July 2008. Plant parts were air-dried at room
temperature in a shady place and kept from direct light.
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Ö. Aksakal, H. Uysal, D.A. Çolak, E. Mete, Y. Kaya MAKÜ FEBED 3(1): 32-36 (2012)
Palandöken Mountain were (E)-2-decenal 52.22%, (2E)dodecenal 15.78%, 8S.14-cedranediol 8.51% and ntetradecenal 5.53%. This results show that E-2-decenal,
(2E)- dodecenal, dodecenal and thymol quantities in the
essential oil composition of Cymbocarpum erythraeum
is higher than those of Cymbocarpum wiedemannini
(Baser & Ozer, 1999). While (E)–2-tetradecanol,
decanoic acid, 2-decanoic acid and 2 dodecanoic acids
exist in high amount in Cymbocarpum wiedemanni, they
were not detected in Cymbocarpum erythraeum which
we used for our experiments. Lastly, n tetradecanol and
8S, 14-cedranediol compounds were high in
Cymbocarpum erythraeum but these compounds were
not detected in Cymbocarpum widemannii (Baser &
Ozer, 1999).
on percentage of survival from larvae and adult
individuals belonging to Drosophila melanogaster. For
this purpose, the same aged larvae (72±4 hr) were
chronically fed with growth medium containing different
concentrations of CEEO (15, 30, 60, 90, 120 and 150
µL). 100 larvae were used at all the application groups.
In control and control+tween groups, 98 and 95 of the
total number of 100 larvae were found to be matured,
respectively. The difference between control and
control+tween groups on percentage of survival was not
statistically significant (p>0.05). However, in all
application groups, the survival rate showed a significant
decrease depending on the concentration of
Cymbocarpum erythraeum essential oils (Table 2).
In this study, the effects of essential oils obtained from
Cymbocarpum erythraeum (CEEO) were investigated
Table 1. Chemical composition (expressed as %) of
Cymbocarpum erythraeum essential oil
Components
RI*
Composition (%)
n-Octanal
998
0.48
(Z)-Linalol oxide (Furanoid)
1067
0.43
(E)- linalol oxide (Furanoid)
1094
0.47
n-Nonanal (Pelargonaldehyde)
1100
0.42
(2E)-Nonen-1-al
1157
0.23
Naptnalene
1178
0.38
(4Z)-Decanal
1193
1.42
(E)-4-Decenal
1196
5.08
(3E)-Decen-2-one
1234
0.97
(E)-2-Decenal
1260
52.22
Thymol
1289
0.38
(8Z)-Undecanal
1300
0.35
Undecanal
1305
0.29
Piperitenone oxide
1366
0.42
n-undecanal
1367
0.35
3-Dodecanone
1389
0.74
Dodecanal (Lauraldehyde)
1408
1.41
(2E)-Dodecenal
1464
15.78
Tetradecanal (Myristaldehyde)
1611
0.25
n-Tetradecanol
1671
5.53
8S,14-Cedranediol
1889
8.51
n-Eicosane
2000
0.25
Total
96.36
*
RI, retention index; compound listed in order of elution from a BPX5
MS column; identification: GC-MS, RI
Table 2. Larval lethality and survival rates with chronic treatment to adult flies of Cymbocarpum
erythraeum essential oil
CEEO (µL)
Letalite
Survival
Female Survival
Male Survival
Application Groups
Total Larvae
(%)
(%)
(%)
(%)
Control
100
0,02
0,98a
0,51
0,47*
Control+Tween
100
0,05
0,95a
0,47
0,48*
100
0,79
0,21b
0,10
0,11
15 µL
100
0,94
0,06c
0,02
0,03
30 µL
100
0,97
0,03d
0,01
0,02
60 µL
100
90 µL
100
120 µL
100
150 µL
a-d
: Values in the same line showed significant different at %5 level, *: Values in the same line showed is not
significant
34
100
80
60
40
20
15
0µ
L
12
0µ
L
90
µL
60
µL
30
µL
15
µL
0
C
on
tro
C
on
l
tro
l+
Tw
ee
n
percentage of total survival (%)
Ö. Aksakal, H. Uysal, D.A. Çolak, E. Mete, Y. Kaya MAKÜ FEBED 3(1): 32-36 (2012)
Concentrations
100
80
percentage of male
survi val (%)
60
percentage of female
survi val (%)
40
20
L
L
15
0µ
12
0µ
90
µL
60
µL
30
µL
15
µL
0
C
C
o
on
tro ntro
l
l+
Tw
ee
n
percentage of survival (%)
Figure 1. Survival percentages of larvae after chronic feeding
of 72±4 h larvae with Cymbocarpum erythraeum essential oil
of different concentrations
Concentrations
Figure 2. Comparison of percentage survival of male and female adults
had excellent inhibitor effects against D. melanogaster
larvae. Moreover, the lowest 15 µL dose tested was also
toxic, killing 79% larvae, in the controls the survival was
98%.
When the application groups (15, 30 and 60 µL) were
compared with control group, 21 (10♀♀; 11♂♂), 6
(2♀♀; 3♂♂) and only 3 (1♀; 2♂♂) of larvae could be
survived. In the first three application groups (15, 30 and
60 µL), the survival rate of larvae was changing between
21% and 3%. All of the survival adult individuals
matured from larvae to adult survived in 15 µL and 60
µL application groups. Only, 6 of 100 larvae matured in
30 µL application group. But 5 of 6 larvae were able to
survive (Table 2). All of the larvae belonging to 90 µL
application group became pupae but any adult was not
obtained. Even in the 120 µL and 150 µL application
groups, larvae failed to become prepupa (Fig. 1). When
the application groups were compared with control
group, it was found that the difference was statistically
important (p<0.05).
Mortality of larvae exposed to the essential oils may be
the results of many parameters, including the structure,
function and biochemistry of the Drosophila growth in
relation to the developmental time (Reynolds, 1987).
It is well documented that many essential oils and their
constituents affect biochemical process, which
specifically disrupt the endocrinologic balance of insects
(Pavlidou et al., 2004). They may be neurotoxic or may
act as insect growth regulators, disrupting the normal
process of developmental time (Balandrin & Klocke,
1988). Additionally, when D. melanogaster embryos or
larvae are exposed to different doses of essential oils,
various types of somatic mutation or even mitotic
recombination effects may be induced in a number of
imaginal disk cells (Würgler & Vogelf, 1986).
Observation on the mortality caused by the compounds
used in the present study showed that most of the
larvae of D. melanogaster were chronically poisoned.
Similar studies have shown that plant essential oils
posse significant larvicidal, adulticidal and insecticidal
activities on D. melanogaster. For example, Karpouhtsis
et al., (1998) reported that Oregano essential oils exhibit
good larvicidal effect on D. melanogaster. Similarly,
Pavlidou et al., (2004) found that the essential oils of
Salvia fruticosa and Mentha plegium were toxic against
In control and control+ tween application group, the rate
of male and female was 1:1. There were the least
survival male and female individuals in the first three
application groups. For this reason, it is so impossible to
say about the effects of Cymbocarpum erythraeum
essential oils on rate of male and female (Fig. 2).
In the present study, essential oils obtained from
Cymbocarpum erythraeum were examined for larvicidal
and adulticidal activity against D. melanogaster.
According to GC-MS analyses, the major constituents of
the essential oil of the aerial parts in this plant were (E)2-decenal and (2E)-dodecenal. Results obtained from
the larvicidal test using the essential oil from this plant
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Ö. Aksakal, H. Uysal, D.A. Çolak, E. Mete, Y. Kaya MAKÜ FEBED 3(1): 32-36 (2012)
Karpouhtsis, I., Pardali, E., Feggou, E., Kokkini, S.,
Scouras, Z.G., Mavragani-Tsipidou, P. (1988).
Insecticidal and genotoxic activities of oregano
essential oils”. J Agric Food Chem, 46(3), 11111115.
Kim, J., Seo, S.M., Lee, S.G., Shin, S.C., Park, I.K.
(2008). Nematicidal activity of plant essential oils
and components from Coriander (Coriandrum
sativum), Oriental sweetgum (Liquidambar
orientalis), and Valerian (Valeriana wallichii)
essential oils against pine wood Nematode
(Bursaphelenchus xylophilus). J Agric Food
Chem, 56(16), 7316-7320.
Miyazawa, M., Tsukamoto, T., Anzai, J., Ishikawa, Y.
(2004). Insecticidal Effect of Phthalides and
Furanocoumarins from Angelica acutiloba against
Drosophila melanogaster. J Agric Food Chem,
52(14), 4401-4405.
Pavlidou, V., Karpouhtsis, I., Franzios, G., Zambetaki,
A., Scouras, Z., Mavragani-Tsipidou, P. (2004).
Insecticidal and genotoxic effects of essential oils
of Greek sage, Salvia fruticosa, and Mint, Mentha
pulegium, on D. melanogaster and Bactrocera
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that essential oil of Coriandrum sativum which contains
especially
(E)-2-decenal,
possess
significant
nematocidal and insecticidal activities. According to our
results, (E)-2-Decenal (52.22%) is the most abundant
compound in the essential oils of Cymbocarpum
erythraeum (Table 1). So this compound leads to
larvicidal activity on D. melanogaster as its nematocidal
and insecticidal activities. The excellent larvicidal
activities of the essential oils of Cymbocarpum
erythraeum demonstrated us that it has natural
insecticides potentiality.
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